Figure 2.

Evofosfamide‐induced apoptosis in breast cancer cells in vitro. (A) Four breast cancer cell lines were seeded in 96‐well plates at 1 × 10⁴ cells per well and treated with evofosfamideat 50 μmol/L or coincubated with the broad specificity caspase inhibitor z‐VAD‐fmk (50 μmol/L). To exclude possible toxic effects of the inhibitor, cells were also treated with the inhibitor alone under normoxic and hypoxic (1% O₂) conditions. Cell lysates were used to determine caspase‐3‐like activity, using the caspase‐3‐specific fluorogenic substrate, zDEVD‐AFC, and cell viability was assessed via crystal violet staining. Data points show means of quadruplicate results from a representative experiment, repeated at least twice; bars ± SD.